阅读说明：本技术 一种红薯中天然生物胶的制备方法及应用 (Preparation method and application of natural biogel in sweet potatoes ) 是由 舒友菊 刘锦琼 薛冬 王自力 刘萍 贾晓慧 王青云 郭永仓 于 2021-09-17 设计创作，主要内容包括：本发明涉及一种红薯中天然生物胶的制备方法及应用,将新鲜洗干净的红薯用捣碎机制成红薯粉末,取一定量红薯粉末加入水中,搅拌后反复过滤,得红薯渣和滤液；将滤液静置后取上清液至灭菌容器中,同时加入红薯渣,混合搅拌后加入纤维素水解酶酶解3h,再加入木瓜蛋白酶酶解2h；之后加入淀粉酶酶解1h,过滤后的滤液加入海藻酸钠,搅拌后放到65-80℃水浴锅中搅拌反应10-15h；所得混合液冷冻干燥,即得天然生物胶。本发明以红薯为原料,利用生物酶解法获得生物活性物质,与海藻酸钠反应制备天然生物胶,该生物胶对癌细胞具有显著的抑制作用,为红薯中多肽、多糖、黄酮和果胶的开发提供了一定的理论依据,为以后产业开发及产业升级奠定了基础。(The invention relates to a preparation method and application of natural biological glue in sweet potatoes, which comprises the steps of preparing sweet potato powder from fresh and clean sweet potatoes by a pounding machine, adding a certain amount of sweet potato powder into water, stirring and repeatedly filtering to obtain sweet potato residues and filtrate; standing the filtrate, taking supernatant into a sterilization container, simultaneously adding sweet potato residue, mixing and stirring, adding cellulose hydrolase for enzymolysis for 3h, and adding papain for enzymolysis for 2 h; adding amylase for enzymolysis for 1h, adding sodium alginate into the filtered filtrate, stirring, and placing into a 65-80 ℃ water bath kettle for stirring reaction for 10-15 h; and freeze-drying the obtained mixed solution to obtain the natural biological glue. The invention takes sweet potato as raw material, obtains bioactive substance by biological enzymolysis method, and reacts with sodium alginate to prepare natural biological glue, which has significant inhibiting effect on cancer cells, provides certain theoretical basis for the development of polypeptide, polysaccharide, flavone and pectin in sweet potato, and lays foundation for the future industrial development and industrial upgrading.)

1. A preparation method of natural biogel in sweet potatoes is characterized by comprising the following steps:

(1) separating sweet potato starch:

preparing fresh and clean sweet potatoes into sweet potato powder by using a pounding machine, adding a certain amount of the obtained sweet potato powder into water, homogenizing and stirring the formed mixed system for 30 minutes at normal temperature, repeatedly filtering the mixed system by using gauze until the filtered liquid is colorless and transparent, and finally obtaining sweet potato residues and filtrate.

(2) Preparing sweet potato polypeptide:

standing the filtrate obtained in the step (1) at normal temperature for 60 minutes, transferring the supernatant into a sterilized container, adding the sweet potato residue obtained in the step (1) into the sterilized container, stirring the mixed system at a constant speed for 20 minutes, adding cellulose hydrolase, adjusting the pH of the mixed system to about 9.0, and performing enzymolysis for 3 hours under stirring at normal temperature; then adding papain, adjusting the pH value of the reaction system to about 9.0, and carrying out enzymolysis for 2 hours under stirring at normal temperature;

(3) sweet potato residue starch hydrolysis

Adding amylase into the reaction system obtained after enzymolysis of the papain in the step (2), adjusting the pH value of the obtained mixed system to be about 8.0, carrying out enzymolysis for 1h under stirring at normal temperature, filtering, and removing filter residues to obtain a filtrate;

(4) preparation of sweet potato natural biological glue

Adding sodium alginate into the filtrate obtained in the step (3), stirring for 30 minutes, putting into a 65-80 ℃ water bath kettle, adjusting the pH value of a reaction system to be about 7.0, and reacting in the water bath kettle for 10-15 hours under stirring; and freeze-drying the obtained mixed solution to obtain the natural biological glue.

2. The method for preparing natural biogel in sweet potatoes as claimed in claim 1, wherein the volume ratio of solid to liquid in a mixing system formed after the sweet potato powder is added into water in the step (1) is 1: 500.

3. The method for preparing natural biogel in sweet potatoes as claimed in claim 1 or 2, wherein the mass concentration of cellulose hydrolase in the step (2) is 0.2-0.6%, and the ratio of the addition volume of the cellulose hydrolase to the mass of the sweet potato powder used in the step (1) is 1ml:100 g.

4. The method for preparing natural biogel in sweet potatoes as claimed in claim 1, wherein the mass ratio of the added mass of the papain in the step (2) to the mass of the sweet potato residue obtained in the step (1) is 1: 100000.

5. The method for preparing natural biogel in sweet potatoes as claimed in claim 1, wherein the mass concentration of the amylase in the step (3) is 0.2%, and the ratio of the addition volume of the amylase to the mass of the sweet potato powder used in the step (1) is 2ml:100 g.

6. The method for preparing natural biogel in sweet potatoes as claimed in claim 1, wherein the mass ratio of the added mass of the sodium alginate in the step (4) to the mass of the sweet potato powder used in the step (1) is 1: 500.

7. The method for preparing natural biogel in sweet potatoes as claimed in claim 1, wherein the reagent for adjusting the pH is sodium bicarbonate solution or sodium hydroxide solution.

8. Application of natural biogel in sweet potato in treating and preventing cancer is provided.

9. Application of natural biogel in sweet potato in treating and preventing liver cancer is provided.

Technical Field

The invention relates to the technical field of biology, and particularly relates to a preparation method and application of natural biogum in sweet potatoes.

Background

Sweet potatoes (Ipomoea batatas (L.)) are native to Central America, are Convolvulaceae plants of Tulipae, and are named sweet potatoes, and other sweet potatoes, sweet potatoes and the like are commonly used. The Chinese characters are introduced from the early stage of the Ming Dynasty perpetual calendar, and the Chinese characters have profound influence on agriculture and economy in China. According to FAO (world food and agricultural organization) statistics, the total yield of Chinese sweet potatoes is 5.20 multiplied by 10 in 2019⁷Ton, harvest area 2.37X 10⁶Hectare, 2.19 tons per hectare. Currently, sweet potatoes remain one of the most prominent food crops.

Scientific research shows that each 100g of sweet potatoes contains 1.8g of protein, 29.5g of sugar, 0.2g of fat, 20mg of phosphorus, 18mg of calcium, 0.4mg of iron and 1.1mg of carotene, and also contains nicotinic acid, vitamins, brass, pectin, linoleic acid, dietary fibers, lysine, polysaccharide, polypeptide and the like, wherein the content of the vitamins is 3-6 times higher than that of normal grains, and the vitamins contain lysine which is richer than the grains.

Sweet potato polypeptide, polysaccharide, flavone, pectin and the like have strong biological activity, certain water solubility and high nutritional value, and can be applied to the industries of medical care and food processing. Therefore, the active substances such as polypeptide, polysaccharide, flavone and pectin in the sweet potatoes and sodium alginate are used for preparing natural colloid for development and serving as future health care products and nutritional products, and the method is beneficial to the upgrading of the sweet potato industry and the increase of the additional value of the sweet potatoes.

Disclosure of Invention

The invention aims to provide a preparation method and application of natural biogel in sweet potatoes. The invention provides a certain theoretical basis for the development of extracting polypeptide, polysaccharide, flavone and pectin mixed substances in sweet potatoes, and lays a foundation for the future industrial development and industrial upgrading.

The invention is realized by the following technical scheme, and the preparation method of the natural biogel in the sweet potatoes provided by the invention specifically comprises the following steps:

(1) separating sweet potato starch:

preparing fresh and clean sweet potatoes into sweet potato powder by using a pounding machine, adding a certain amount of the obtained sweet potato powder into water, homogenizing and stirring the formed mixed system for 30 minutes at normal temperature, repeatedly filtering the mixed system by using gauze until the filtered liquid is colorless and transparent, and finally obtaining sweet potato residues and filtrate.

(2) Preparing sweet potato polypeptide:

standing the filtrate obtained in the step (1) at normal temperature for 60 minutes, transferring the supernatant into a sterilized container, adding the sweet potato residue obtained in the step (1) into the sterilized container, stirring the mixed system at a constant speed for 20 minutes, adding cellulose hydrolase, adjusting the pH of the mixed system to about 9.0, and performing enzymolysis for 3 hours under stirring at normal temperature; then adding papain, adjusting the pH value of the reaction system to about 9.0, and carrying out enzymolysis for 2 hours under stirring at normal temperature;

(3) sweet potato residue starch hydrolysis

Adding amylase into the reaction system obtained after enzymolysis of the papain in the step (2), adjusting the pH value of the obtained mixed system to be about 8.0, carrying out enzymolysis for 1h under stirring at normal temperature, filtering, and removing filter residues to obtain a filtrate;

(4) preparation of sweet potato natural biological glue

Adding sodium alginate into the filtrate obtained in the step (3), stirring for 30 minutes, putting into a 65-80 ℃ water bath kettle, adjusting the pH value of a reaction system to be about 7.0, and reacting in the water bath kettle for 10-15 hours under stirring; and freeze-drying the obtained mixed solution to obtain the natural biological glue.

Further, after the sweet potato powder in the step (1) is added into water, the solid-liquid volume ratio of the formed mixed system is 1: 500.

Further, the mass concentration of the cellulose hydrolase in the step (2) is 0.2-0.6%, and the ratio of the added volume of the cellulose hydrolase to the mass of the sweet potato powder used in the step (1) is 1ml:100 g.

Further, the mass ratio of the added mass of the papain in the step (2) to the mass of the sweet potato residue obtained in the step (1) is 1: 100000.

Further, the mass concentration of the amylase in the step (3) is 0.2%, and the ratio of the addition volume of the amylase to the mass of the sweet potato powder used in the step (1) is 2ml:100 g.

Further, the ratio of the added mass of sodium alginate in the step (4) to the mass of the sweet potato powder used in the step (1) is 1: 500.

Further, the reagent used for adjusting the pH is sodium bicarbonate solution or sodium hydroxide solution.

The natural biogel in the sweet potatoes obtained by the invention can be applied to the treatment and prevention of cancers, in particular to the treatment and prevention of liver cancers.

The biological enzymolysis technology makes the substances fully decomposed by the action of enzyme to obtain the extract required by the biological engineering. Meanwhile, the characteristics of different active ingredients of the substance can be mastered by utilizing different catalytic actions of different enzymes, so that the active substance can be extracted. The biological enzyme is a special biological catalyst which is generated by living cells of organisms and exists in the form of protein, can decompose plant cell walls under the conditions of normal temperature, normal pressure and mild acid and alkali, greatly improves the extraction rate of effective components in natural plants, improves the filtration speed and purification effect in the production process, and improves the product purity and the quality of a preparation.

Compared with the prior art, the invention has the following advantages:

the invention takes sweet potato as raw material, uses cellulose hydrolase to dissolve out flavone, pectin and a small part of polypeptide in the sweet potato, adds papain to extract polypeptide substance, adds amylase to hydrolyze to obtain polysaccharide, and finally mixes the flavone, pectin, polypeptide and polysaccharide in the filtrate with sodium alginate to react to obtain the biological glue. The biological glue has obvious inhibition effect on liver cancer cells, and is a natural anticancer biological glue. The invention utilizes a biological enzymolysis method to extract polysaccharide, flavone, polypeptide and pectin in sweet potatoes, and the polysaccharide, the flavone, the polypeptide and the pectin react with sodium alginate to prepare biological glue, the reaction condition is mild, higher natural biological activity is reserved, the enzymolysis rate of protein and cellulose of the sweet potatoes reaches 90-100%, cheap raw materials are provided for biological health industry, high-valued utilization of the sweet potatoes is realized, the requirements on process equipment are not strict, and the industrial application is easy.

Drawings

FIG. 1 is an SEM image of a natural biological glue obtained by the present invention;

FIG. 2 shows the inhibition effect of the natural biogel on liver cancer cells in different energies;

FIG. 3 shows the inhibition of the natural biogel on liver cancer cells at different times.

Detailed Description

For a better understanding of the contents of the invention, reference will now be made to the following examples and accompanying drawings which illustrate the invention. The present embodiment is implemented based on the technology of the present invention, and a detailed implementation manner and operation steps are given, but the scope of the present invention is not limited to the following embodiments.

The invention relates to a preparation method of natural biogel in sweet potatoes, which specifically comprises the following steps:

(1) separating sweet potato starch:

preparing fresh and clean sweet potatoes into sweet potato powder by using a pounding machine, adding the sweet potato powder into water, and forming a mixed system with the solid-liquid volume ratio of 1: 500; homogenizing and stirring at normal temperature for 30 min, and repeatedly filtering with gauze until the filtrate is colorless and transparent to obtain sweet potato residue and filtrate.

(2) Preparing sweet potato polypeptide:

standing the filtrate obtained in the step (1) at normal temperature for 60 minutes, transferring the supernatant into a sterilized container, adding the sweet potato residue obtained in the step (1) into the sterilized container, uniformly stirring the mixed system for 20 minutes, adding cellulose hydrolase with the mass concentration of 0.2-0.6%, adjusting the pH of the mixed system to about 9.0, and carrying out enzymolysis for 3 hours under normal-temperature stirring; then adding papain into a reaction system, adjusting the pH value to be about 9.0, and carrying out enzymolysis for 2 hours under stirring at normal temperature; wherein the ratio of the adding volume of the cellulose hydrolase to the mass of the sweet potato powder used in the step (1) is 1ml:100 g; the mass ratio of the added mass of the papain to the mass of the sweet potato residue obtained in the step (1) is 1: 100000.

(3) sweet potato residue starch hydrolysis

Adding amylase with the mass concentration of 0.2% into the reaction system obtained after the papain is subjected to enzymolysis in the step (2), adjusting the pH value of the obtained mixed system to be about 8.0, performing enzymolysis for 1 hour under stirring at normal temperature, filtering, and removing filter residues to obtain filtrate; wherein the mass ratio of the added volume of the amylase to the sweet potato powder used in the step (1) is 2ml to 100 g.

(4) Preparation of sweet potato natural biological glue

Adding a certain amount of sodium alginate into the filtrate obtained in the step (3), stirring for 30 minutes, putting into a 65-80 ℃ water bath kettle, adjusting the pH value of a reaction system to be about 7.0, and reacting in the water bath kettle for 10-15 hours under stirring; placing the obtained mixed solution in a freeze dryer, and freeze-drying to obtain the natural anticancer biological glue. Wherein the mass ratio of the added sodium alginate to the sweet potato powder used in the step (1) is 1: 500.

The pH can be adjusted in the above process with sodium bicarbonate solution or sodium hydroxide solution.

According to the invention, after the cellulose hydrolase is added in the step (2), flavone, pectin and a small part of polypeptide in sweet potatoes can be dissolved out, the papain is added to extract polypeptide substances, after the amylase is added in the step (3), the starch is hydrolyzed to obtain polysaccharide, and finally the flavone, the pectin, the polypeptide and the polysaccharide in the filtrate are mixed with the sodium alginate to react to obtain the biological colloid. In the method, the enzymolysis rate of starch, protein and cellulose of sweet potatoes can reach 90-100%.

The following is a detailed description of specific embodiments.

Example 1:

(1) preparing fresh and clean sweet potatoes into sweet potato powder by using a pounding machine, adding 500g of sweet potato powder into water, and forming a mixed system with the solid-liquid volume ratio of 1: 500; after stirring homogeneously at room temperature for 30 minutes, the mixture was repeatedly filtered with gauze until the filtrate was colorless and transparent, and about 285g of sweet potato residue and filtrate were obtained.

(2) Preparing sweet potato polypeptide:

standing the filtrate obtained in the step (1) at normal temperature for 60 minutes, transferring the supernatant into a sterilized container, adding the sweet potato residue obtained in the step (1) into the sterilized container, uniformly stirring the mixed system for 20 minutes, then adding 5ml of cellulose hydrolase with the mass concentration of 0.2%, adjusting the pH of the mixed system to be about 9.0, and carrying out enzymolysis for 3 hours under normal-temperature stirring; then adding papain into a reaction system, adjusting the pH value to be about 9.0, and carrying out enzymolysis for 2 hours under stirring at normal temperature; the mass ratio of the added mass of the papain to the mass of the sweet potato residue obtained in the step (1) is 1: 100000.

(3) sweet potato residue starch hydrolysis

Adding 10ml of amylase with the mass concentration of 0.2% into the reaction system obtained after the papain is subjected to enzymolysis in the step (2), adjusting the pH value of the obtained mixed system to be about 8.0, performing enzymolysis for 1 hour under stirring at normal temperature, filtering, and removing filter residues to obtain a filtrate;

(4) preparation of sweet potato natural biological glue

Adding 1g of sodium alginate into the filtrate obtained in the step (3), stirring for 30 minutes, putting the mixture into a 65 ℃ water bath kettle, adjusting the pH value of a reaction system to be about 7.0, and reacting in the water bath kettle for 15 hours under stirring; and placing the obtained mixed solution in a freeze dryer, and freeze-drying to obtain the biological glue.

Example 2:

(1) preparing fresh and clean sweet potatoes into sweet potato powder by using a pounding machine, adding 500g of sweet potato powder into water, and forming a mixed system with the solid-liquid volume ratio of 1: 500; after stirring homogeneously at room temperature for 30 minutes, the mixture was repeatedly filtered with gauze until the filtrate was colorless and transparent, and about 285g of sweet potato residue and filtrate were obtained.

(2) Preparing sweet potato polypeptide:

standing the filtrate obtained in the step (1) at normal temperature for 60 minutes, transferring the supernatant into a sterilized container, adding the sweet potato residue obtained in the step (1) into the sterilized container, uniformly stirring the mixed system for 20 minutes, then adding 5ml of cellulose hydrolase with the mass concentration of 0.4%, adjusting the pH of the mixed system to be about 9.0, and carrying out enzymolysis for 3 hours under normal-temperature stirring; then adding papain into a reaction system, adjusting the pH value to be about 9.0, and carrying out enzymolysis for 2 hours under stirring at normal temperature; the mass ratio of the added mass of the papain to the mass of the sweet potato residue obtained in the step (1) is 1: 100000.

(3) sweet potato residue starch hydrolysis

Adding 10ml of amylase with the mass concentration of 0.2% into the reaction system obtained after the papain is subjected to enzymolysis in the step (2), adjusting the pH value of the obtained mixed system to be about 8.0, performing enzymolysis for 1 hour under stirring at normal temperature, filtering, and removing filter residues to obtain a filtrate;

(4) preparation of sweet potato natural biological glue

Adding 1g of sodium alginate into the filtrate obtained in the step (3), stirring for 30 minutes, putting the mixture into a water bath kettle at the temperature of 75 ℃, adjusting the pH value of a reaction system to be about 7.0, and reacting in the water bath kettle for 12 hours under stirring; and placing the obtained mixed solution in a freeze dryer, and freeze-drying to obtain the biological glue.

Example 3:

(1) preparing fresh and clean sweet potatoes into sweet potato powder by using a pounding machine, adding 500g of sweet potato powder into water, and forming a mixed system with the solid-liquid volume ratio of 1: 500; after stirring homogeneously at room temperature for 30 minutes, the mixture was repeatedly filtered with gauze until the filtrate was colorless and transparent, and about 285g of sweet potato residue and filtrate were obtained.

(2) Preparing sweet potato polypeptide:

standing the filtrate obtained in the step (1) at normal temperature for 60 minutes, transferring the supernatant into a sterilized container, adding the sweet potato residue obtained in the step (1) into the sterilized container, uniformly stirring the mixed system for 20 minutes, then adding 5ml of cellulose hydrolase with the mass concentration of 0.6%, adjusting the pH of the mixed system to about 9.0, and carrying out enzymolysis for 3 hours under normal-temperature stirring; then adding papain into a reaction system, adjusting the pH value to be about 9.0, and carrying out enzymolysis for 2 hours under stirring at normal temperature; the mass ratio of the added mass of the papain to the mass of the sweet potato residue obtained in the step (1) is 1: 100000.

(3) sweet potato residue starch hydrolysis

Adding 10ml of amylase with the mass concentration of 0.2% into the reaction system obtained after the papain is subjected to enzymolysis in the step (2), adjusting the pH value of the obtained mixed system to be about 8.0, performing enzymolysis for 1 hour under stirring at normal temperature, filtering, and removing filter residues to obtain a filtrate;

(4) preparation of sweet potato natural biological glue

Adding 1g of sodium alginate into the filtrate obtained in the step (3), stirring for 30 minutes, putting into a water bath kettle at 80 ℃, adjusting the pH value of a reaction system to be about 7.0, and reacting in the water bath kettle for 10 hours under stirring; and placing the obtained mixed solution in a freeze dryer, and freeze-drying to obtain the biological glue.

FIG. 1 is an SEM image of the natural biological glue obtained by the invention, and the anticancer biological glue of sweet potato has a certain network skeleton structure, certain pores and targeting sites.

The experiment of the inhibition effect of the biological glue on cancer cells comprises the following steps:

(1) in vivo cells of liver cancer patients were cultured for 5 days and divided into 6 groups, the 1 st, 3 rd and 5 th groups were not added with the biogel obtained by the present invention, the 2 nd, 4 th and 6 th groups were added with the same amount of the biogel obtained by the present invention, the 1 st and 2 nd groups grew under 1.00W of light energy, the 3 rd and 4 th groups grew under 1.50W of light energy, the 5 th and 6 th groups grew under 2.00W of light energy, and the change of cancer cells and normal cells in different times was observed, and the results are shown in FIG. 2. Under three different light energies of 1.00W, 1.50W and 2.00W, the first horizontal row of pictures under each light energy is the competition of cancer cells and normal cells when no biological glue is added, and the second horizontal row of pictures is the competition of cancer cells and normal cells with the time after the biological glue obtained by the invention is added. The second horizontal panel shows bright spots indicating that normal cells have resumed growth. As can be seen from FIG. 2, under different light energies, the normal cells in the first row die, and the cancer cells gradually weaken with the passage of time after the biogel obtained by the present invention is added in the second row, which shows that the biogel prepared by the present invention has obvious inhibition effect on liver cancer cells.

(2) Culturing in vivo cells of liver cancer patients for 5 days, dividing into 9 groups, adding different amounts of the biogel obtained by the invention, gradually increasing the adding amount of the biogel from the 1 st group to the 9 th group, growing by using light energy, observing the change of cancer cells and normal cells in different time, wherein the 1 st group is at the leftmost side, and the 9 th group is at the rightmost side, and the result is shown in fig. 3. The appearance of bright spots in the pictures indicates that normal cells restored growth. The addition of the biological glue from left to right is gradually increased, the addition of the biological glue in the first row from the left is less, the cancer cells are not controlled until the sixth day, the addition of the biological glue in the second row has a certain inhibition effect on the cancer cells, and the cancer cells are gradually weakened and the normal cells are restored to grow from left to right along with the increase of the addition of the biological glue.

The above description is only an embodiment of the present invention, and is not intended to limit the present invention in any way, and the present invention may also have other embodiments according to the above structures and functions, and is not listed again. Therefore, any simple modification, equivalent change and modification of the above embodiments according to the technical essence of the present invention by those skilled in the art can be made within the technical scope of the present invention.